Comparison of molecular mutations of G6PD deficiency gene between icteric and nonicteric neonates.

Jaundice is a common disorder in neonates and one of the provable causes of glucose-6-phosphate dehydrogenase (G6PD) deficiency, some mutation types of which may be associated with severe neonatal icter. The present study has been conducted to compare G6PD mutations in incteric and non icteric neonates. This case-control study was implemented in the NICU and Newborn Ward of Amirkola Children Hospital in 2007-2008. Available sampling approach was used and 50 icteric as well as 50 non-icteric newborns, both with G6PD deficiency, were selected as the case and the control group respectively. G6PD deficiency was diagnosed using FST (Fluorescent Spot Test) method. All samples were first evaluated in terms of Mediterranean mutation and the negative cases were then examined for Chatham mutation; all remaining samples were finally tested for Cosenza mutation. G6PD mutations were compared in the two groups and P-value less than 0.05 was considered significant. In icteric group, 76% were male and 24% were female and in non-icteric group, 70% were male and 30% were female. The mean weight of neonates was 3.2 ± 0.4 kg and 2.8 ± 0.8 kg in icteric and non-icteric groups respectively (p<0.05). In non-icteric group, 54% Mediterranean, 18% Chatham, and 28% Cosenza negative were observed and in icteric group, 56% Mediterranean, 32% Chatham, and 12% Cosenza negative were found; the distribution of Mediterranean and Chatham mutations was not significantly different between the two groups (p>0.05), however, the distribution of rare mutations (Cosenza negative) was significantly different between icteric and non-icteric groups with enzyme deficiency (p<0.05). The mean bilirubin level was not statistically different in Mediterranean (18.5±2.9), Chatham (18.8±2.1) and Cosenza negative (20±4.3) mutations (p> 0.05). Newborns with Chatham mutation have been less in need of exchange transfusions (p <0.05) indicating that rare mutations of G6PD gene may less likely lead to neonatal icter.

of this enzyme increases erythrocyte susceptibility to oxidative stress (2). The disorder was first diagnosed in 1956 and many investigations have been conducted since then (3); the Electrophoretic forms of which were identified in 1960 (4).
G6PD deficiency is highly different in terms of diversity and distribution; almost 7.5% of people in the world are the carriers of one or two G6PD deficiency genes (5). In an investigation in the city of Babol, the prevalence of the disorder has been reported to be 12.5% in males and 4.1% in females (5).
Glucose-6-phosphatase gene is located on the distal part of the long arm of chromosome X at Xq28 position (2,5). With 13 exons and 12 introns, the gene has a length of approximately 18.5 kb, encoding a 59 kDa polypeptide with 515 amino acids; the products of this gene form a number of homodimers or tetramers (5,(7)(8)(9).
One of the most common G6PD variants is the Mediterranean type which is often associated with favism (13)(14)(15).

Studies conducted in Iran -in Mazandaran
Province and Sari city -by Mesbah et al., showed that the prevalence of Chatham mutation was higher in this region compared with other parts of the world between three different G6PD polymorphic variants, including 66.2% Mediterranean, 27% Chatham and 6.75% Cosenza (16), and, hence, the prevalence of all G6PD types in this region is more similar to that in Italy in comparison with other Mediterranean countries (16). Regarding the studies performed, Mediterranean G6PD is the most common mutation in Iran as well as other tropical and subtropical regions (5,13,(15)(16)(17).
Neonatal icter is the most common clinical manifestation of G6PD deficiency (18). Icter is developed in one third of newborns with the above-  (19).
Considering G6PD deficiency, especially in the Northern regions, and its association with neonatal icter and related complications, and the fact that no study has been conducted in Iran in terms of the relationship between enzymeassociated specific DNA mutations and clinical manifestations in neonatal age group, the present study aimed to investigate three common G6PD gene mutations in Mazandaran Province in neonatal age group and its association with the incidence and the severity of neonatal icter.

Subjects
This case-control study was implemented in the NICU and Newborn Ward of Shafizadeh

DNA extraction
Alkalin lysis method was used for the extrac-tion of genomic DNA from 500 µl (0.5 ml) of blood.

DNA amplification
For the evaluation of each of the above mutations, mutation-related region in G6PD gene was first amplified for all DNA samples using PCR approach.
To determine the Mediterranean mutation,      (Table 3).

Discussion
The results of the present study demonstrated that Mediterranean mutation was the most frequent mutation in icteric and non-icteric neonates with G6PD deficiency; this mutation has been observed in more than half of the newborns in both groups, between which the relative distribution was not statistically different; although the distribution of Chatham mutation was higher in icteric neonates, the difference was not significant (